This document demonstrates the use of the bRF and LASSO-D3S functions for integrative GRN inference.

Those functions infer the regulatory pathways of Arabidopsis thaliana’s roots in response to nitrate (N) induction from Varala et al., 2018.

They use as inputs the expression profiles of N-responsive genes and TFBS information. Prior TFBS information was built by searching in the promoters of the N-responsive genes the PWM of the N-responsive regulators.

Data import

Import of the expression data and the N-responsive genes and regulators :

load('rdata/inference_input_N_response_varala.rdata')
genes <- input_data$grouped_genes; length(genes)
## [1] 1426
tfs <- input_data$grouped_regressors; length(tfs)
## [1] 201
counts <- input_data$counts; dim(counts)
## [1] 1426   45
load("rdata/pwm_occurrences_N_response_varala.rdata")
dim(pwm_occurrence)
## [1] 1426  201
ALPHAS <- seq(0,1, by = 0.1)

subset <- sample(genes, replace = F, size = 20)
#subset <- genes


load("results/brf_perumtations_mse_all_genes.rdata")


load("results/100_permutations_bRF_importances.rdata")

Is the mse variation worsened by unmapping prior values and expression profiles of TFs?

For Rfs

draw_gene <- function(gene){
  lmses[gene,] %>%
    gather() %>%
    separate(key, into = c("alpha", "rep", "MSEtype"), sep = " ") %>%
    ggplot(aes(x=alpha, y=value, group =interaction(rep, MSEtype), 
               color = MSEtype))+ggtitle(gene)+ylab("MSE/Var(gene)")+
    xlab("alpha")+
    geom_line()+ggtitle(gene)+theme_pubr(legend = "top")
}


draw_gene_mean_sd <- function(gene, title = NULL){
  lmses[gene, ] %>%
    gather() %>%
    separate(key,
             into = c("alpha", "rep", "MSEtype"),
             sep = " ") %>%
    group_by(alpha, MSEtype) %>%
    mutate(mean_mse = mean(value, na.rm = T),
           sd_mse = sd(value, na.rm = T)) %>%
    ggplot(aes(
      x = as.numeric(alpha),
      y = value,
      color = MSEtype,
      fill = MSEtype
    )) +ggtitle(paste(gene, title))+ylab("MSE/Var(gene)")+
    geom_ribbon(aes(ymin = mean_mse - sd_mse , 
                    ymax = mean_mse + sd_mse  ), 
                alpha = .4)  +theme_pubr(legend = "top")+
    geom_point(alpha = 0.1) + geom_smooth(se=F)+xlab("alpha")

}



draw_gene_effective_integration <- function(gene){
  tfs_with_motif <- names(which(pwm_occurrence[gene,]==1))
  data.frame(lapply(mats, function(mat){mean(rank(mat[,gene])[tfs_with_motif])})) %>%
  gather(key = "setting", value = "summed_importance") %>%
  separate(setting, into = c("method", "alpha", "dataset", "rep"), sep = "_") %>%
  group_by(alpha, dataset) %>%
  mutate(mean_imp = mean(summed_importance),
            sd_imp = sd(summed_importance),
         alpha = as.numeric(alpha))%>%
  ggplot(aes(x=alpha, y = summed_importance, color = dataset, fill = dataset)) +
  geom_point(alpha = 0.1) + geom_smooth(se=F)+
  geom_ribbon(aes(ymin = mean_imp - sd_imp , 
                    ymax = mean_imp + sd_imp  ), 
                alpha = .4)  +theme_pubr(legend = "top") + 
    ylab("effective data integration")
}


draw_MSE_vs_gene_effective_integration <- function(gene){
  tfs_with_motif <- names(which(pwm_occurrence[gene,]==1))
  
  inte <- data.frame(lapply(mats, function(mat){mean(rank(mat[,gene])[tfs_with_motif])})) %>%
  gather(key = "setting", value = "summed_importance") %>%
  separate(setting, into = c("method", "alpha", "dataset", "rep"), sep = "_") %>%
  group_by(alpha, dataset) %>%
  mutate(mean_imp = mean(summed_importance),
            sd_imp = sd(summed_importance),
         alpha = as.numeric(alpha))
  lmses[gene, ] %>%
    gather() %>%
    separate(key,
             into = c("alpha", "rep", "MSEtype"),
             sep = " ") %>%
    group_by(alpha, MSEtype) %>%
    rename(dataset = MSEtype)%>%
    mutate(mean_mse = mean(value, na.rm = T),
           sd_mse = sd(value, na.rm = T),
           alpha = as.numeric(alpha),
           dataset = str_replace(dataset, "true_data", "trueData")) %>%
    full_join(inte, by = c("alpha", "dataset", "rep")) %>%
    group_by(dataset) %>%
    dplyr::arrange(summed_importance) %>% 
    mutate(lower = loess.sd(x=summed_importance, y=value)$lower,
           upper = loess.sd(x=summed_importance, y=value)$upper) %>%
    mutate(lower = frollmean(lower, 1, fill = NA),
           upper = frollmean(upper, 1, fill = NA)) %>%
  ggplot(aes(y=value, x = summed_importance, color = dataset, fill = dataset))+
  geom_ribbon(aes(ymin =lower , 
                  ymax = upper  ), 
                alpha = .4)+ 
  geom_point(alpha = 0.5, size = 0.5)+theme_pubr(legend = "top") + geom_smooth(method = "loess", se = F)+
    ylab("MSE/Var(gene)") + xlab("effective integration") 
}


# 

# 
# loess <- plot(all_data$summed_importance,
#               loess.sd(all_data$summed_importance, all_data$value)$sd)
# 

for(gene in subset){
    print(draw_gene_effective_integration(gene)+draw_gene_mean_sd(gene)+draw_MSE_vs_gene_effective_integration(gene))
}

# 
# gene <- "AT3G49780"
# bad_rf_rank <- draw_gene_effective_integration(gene)+draw_gene_mean_sd(gene)+draw_MSE_vs_gene_effective_integration(gene)
# gene <- "AT3G17609"
# good_rf_rank <- draw_gene_effective_integration(gene)+draw_gene_mean_sd(gene)+draw_MSE_vs_gene_effective_integration(gene)
# 
# 
# bad_rf_rank/bad_rf_imp
# good_rf_rank/good_rf_imp

On the class of interest:

load("results/clusters_mse_bRF_100permutations.rdata")
pos_genes <- names(which(clusters_rf==2))
for(gene in sample(pos_genes,10, replace = F)){
    print(draw_gene_effective_integration(gene)+draw_gene_mean_sd(gene)+draw_MSE_vs_gene_effective_integration(gene))
}

lasso

load("results/lasso_perumtations_mse_all_genes.rdata")
load("results/100_permutations_lasso_importances_freq.rdata")

for(gene in subset){
    print(draw_gene_effective_integration(gene)+draw_gene_mean_sd(gene)+draw_MSE_vs_gene_effective_integration(gene))
}

On the class of interest :

load("results/clusters_mse_lasso_100permutations.rdata")
pos_genes <- names(which(clusters_lasso==1))
for(gene in sample(pos_genes,20, replace = F)){
    print(draw_gene_effective_integration(gene)+draw_gene_mean_sd(gene)+draw_MSE_vs_gene_effective_integration(gene))
}